Cholesterol and other lipids are transported in body fluids by low-density lipoproteins (LDL) and high-density lipoproteins (HDL). Substances that effectuate mechanisms for lowering LDL-cholesterol may serve as effective antihypercholesterolemic agents because LDL levels are positively correlated with the risk of coronary artery disease.
MEVACOR (lovastatin; mevinolin) and ZOCOR (simvastatin) are members of a group of active antihypercholesterolemic agents that function by inhibiting the rate-limiting step in cellular cholesterol biosynthesis, namely the conversion of hydroxymethylglutarylcoenzyme A (HMG-CoA) into mevalonate by HMG-CoA reductase.
The general biosynthetic pathway of a naturally occurring HMG-CoA reductase inhibitor has been outlined by Moore, et al., who showed that the biosynthesis of mevinolin (lovastatin) by Aspergillus terreus ATCC 20542 begins with acetate and proceeds via a polyketide pathway (R. N. Moore, et al., J. Amer. Chem. Soc. 107:3694-3701, 1985). Endo, et al. described similar biosynthetic pathways in Pencillium citrinum NRRL 8082 and Monascus ruber M-4681 (A. Y. Endo, et al., J. Antibiot. 38:444-448, 1985).
The recent commercial introduction of microbial HMG-CoA reductase inhibitors has fostered a need for high yielding production processes. Methods of improving process yield have included scaling up the process, improving the culture medium and simplifying the isolation.
Previous attempts to increase the biosynthesis of HMG-CoA reductase inhibitors at the level of gene expression have focused on increasing the concentration triol polyketide synthase (TPKS), a multifunctional protein with at least six activities as evidenced by the product of the enzymatic activity (Moore, supra, 1985). TPKS is believed to be the rate-limiting enzymatic activity(ies) in the biosynthesis of the HMG-CoA reductase inhibitor compounds.
U.S. Pat. No. 5,744,350 identifies a DNA encoding triol polyketide synthase (TPKS) from Aspergillus terreus. “NPKS” is now preferred to TPKS as the acronym for “nonaketide polyketide synthase.”